Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Gene Expression, Ubiquitin Proteomics, Amplification

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: Stellate cells, LSECs, Kupffer cells and hepatocytes were isolated from the liver of untreated WT mice. WT mice were overdosed with APAP for 1, 6 or 24 hours or fed a CHFD (n=9) for 8 weeks. Panx1 RNA levels were measured in liver cells or tissue. (A) Panx1 RNA in stellate cells, LSECs, Kupffer cells and hepatocytes. (B) Panx1 RNA in liver tissue of WT CHFD and WT APAP mice. Data are expressed as means ± SEM with **p<0.01 and ****p<0.0001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Isolation

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were overdosed with APAP for 1, 6, 24 and 48 hours. (A) Necrotic areas were calculated for WT untreated, WT APAP, Panx1−/− untreated and Panx1−/− APAP mice. (B) Serum ALT and AST activity levels. Data are expressed as means ± SEM with *p<0.05, **p<0.01, ***p<0.001 and ****p<0.0001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Activity Assay

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were fed a CHFD for 8 weeks. (A) Steatosis, lobular inflammation, ballooning score and NAS based on hematoxylin-eosin staining of liver tissue of WT ND, WT CHFD, Panx1−/− ND and Panx1−/− CHFD mice. (B) Serum ALT and AST activity levels. Data are expressed as means ± SEM with *p<0.05, ***p<0.001 and ****p<0.0001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Staining, Activity Assay

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were overdosed with APAP for 1, 6 and 24 hours. (A) APAP-CYS protein adducts were quantified by high-pressure liquid chromatography with electrochemical detection using total liver homogenate. (B) Immunoblot analysis of Cyp2e1 (53 kDa) after separation and blotting, followed by normalization to total protein and expressed as fold change against normalized content of WT mice. (C) Liver GSH and GSSG levels. Data are expressed as means ± SEM with *p<0.05 and **p<0.01.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: High Performance Liquid Chromatography, Western Blot

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were fed a CHFD for 8 weeks. (A) GR, GPx, catalase and SOD activity levels were measured. (B) Immunoblot analysis of SOD (35 kDa), GR (55 kDa), GPx (22 kDa) and catalase (59 kDa) after separation and blotting, followed by normalization to total protein and expressed as fold change against normalized content of WT ND mice. Data are expressed as means ± SEM with *p<0.05, **p<0.01 and ***p<0.001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Activity Assay, Western Blot

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were fed a CHFD for 8 weeks. Serum and liver triglycerides and cholesterol levels were measured. Data are expressed as means ± SEM with **p<0.01 and ***p<0.001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques:

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were overdosed with APAP for 24 hours. Antibody array blots of protein extracts from liver tissue of WT untreated, WT APAP, Panx1−/− untreated and Panx1−/− APAP mice. Data were analyzed by densitometric analysis and were normalized to the average of the positive controls. Data are expressed as means ± SEM with **p<0.01. (BLC, B lymphocyte chemoattractant; CD30L, cluster of differentiation 30 ligand; GCSF, granulocyte colony-stimulating factor; (G)MCSF, (granulocyte)-macrophage colony-stimulating factor; IFN, interferon; IL, interleukin, I-TAC, interferon-inducible T-cell α chemoattractant; MCP, monocyte chemoattractant protein; MIG, monokine induced by interferon-γ; MIP, macrophage inflammatory protein; SDF, stromal cell-derived factor; sTNFR, soluble tumor necrosis factor receptor; TCA, T-cell activation protein; TECK, thymus-expressed chemokine; TIMP, metalloproteinase inhibitor).

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Ab Array, Derivative Assay, Activation Assay

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were fed a CHFD for 8 weeks. Antibody array blots of protein extracts from liver tissue of WT ND, WT CHFD, Panx1−/− ND and Panx1−/− CHFD mice. Data were analyzed by densitometric analysis and were normalized to the average of the positive controls. Data are expressed as means ± SEM with *p<0.05, **p<0.01 and ***p<0.001. (BLC, B lymphocyte chemoattractant; CD30L, cluster of differentiation 30 ligand; GCSF, granulocyte colony-stimulating factor; (G)MCSF, (granulocyte)-macrophage colony-stimulating factor; IFN, interferon; IL, interleukin, I-TAC, interferon-inducible T-cell α chemoattractant; MCP, monocyte chemoattractant protein; MIG, monokine induced by interferon-γ; MIP, macrophage inflammatory protein; SDF, stromal cell-derived factor; sTNFR, soluble tumor necrosis factor receptor; TCA, T-cell activation protein; TECK, thymus-expressed chemokine; TIMP, metalloproteinase inhibitor).

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Ab Array, Derivative Assay, Activation Assay

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were overdosed with APAP for 24 hours. (A) The number of upregulated and downregulated genes represent affected genes in Panx1−/− mice in comparison with WT counterparts. (B) Gene expression levels of Cd36, Cd68 and Cxcl9. Data are expressed as means ± SEM with *p<0.05, **p<0.01 and ***p<0.001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Comparison, Gene Expression

Journal: Biochimica et biophysica acta

Article Title: Protective effect of genetic deletion of pannexin1 in experimental mouse models of acute and chronic liver disease

doi: 10.1016/j.bbadis.2017.12.013

Figure Lengend Snippet: WT and Panx1−/− mice were fed a CHFD for 8 weeks. (A) The number of upregulated and downregulated genes represent affected genes in Panx1−/− mice in comparison with WT counterparts. (B) Gene expression levels of Cd36, Cd68 and Cxcl9. Data are expressed as means ± SEM with *p<0.05, **p<0.01 and ***p<0.001.

Article Snippet: Relative alterations (fold change) in RNA levels were calculated according to the Livak 2 −ΔΔCq formula [ 36 ]. table ft1 table-wrap mode="anchored" t5 caption a7 Gene Assay ID Accession number Assay location Amplicon size (base pairs) Exon boundary Panx1 Mm00450900_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019482.2","term_id":"86262133","term_text":"NM_019482.2"}} NM_019482.2 986 70 3–4 Cd36 Mm00432403_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001159555.1","term_id":"227116346","term_text":"NM_001159555.1"}} NM_001159555.1 1214 104 8–9 Cd68 Mm03047343_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_001291058.1","term_id":"597508191","term_text":"NM_001291058.1"}} NM_001291058.1 956 109 5–6 Cxcl9 Mm00434946_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008599.4","term_id":"162287427","term_text":"NM_008599.4"}} NM_008599.4 243 64 2–6 Actb Mm00607939_s1 {"type":"entrez-nucleotide","attrs":{"text":"NM_007393.3","term_id":"145966868","term_text":"NM_007393.3"}} NM_007393.3 1233 115 6–6 B2m Mm00437762_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_009735.3","term_id":"144227219","term_text":"NM_009735.3"}} NM_009735.3 111 77 1–2 Gapdh Mm99999915_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_008084.2","term_id":"126012538","term_text":"NM_008084.2"}} NM_008084.2 265 107 2–3 Hmbs Mm01143545_m1 {"type":"entrez-nucleotide","attrs":{"text":"NM_013551.2","term_id":"159110430","term_text":"NM_013551.2"}} NM_013551.2 473 81 6–7 Ubc Mm02525934_g1 {"type":"entrez-nucleotide","attrs":{"text":"NM_019639.4","term_id":"157671922","term_text":"NM_019639.4"}} NM_019639.4 370 176 2–2 Open in a separate window caption a8 Primers used for gene expression analysis ( Actb , actin β; B2m , β-2-microglobulin; Gapdh , glyceraldehyde 3-phosphate dehydrogenase; Hmbs , hydroxymethylbilane synthase; Panx1 , pannexin1; Ubc , ubiquitin C).

Techniques: Comparison, Gene Expression

Journal: Biology Methods & Protocols

Article Title: PrimerPooler: automated primer pooling to prepare library for targeted sequencing

doi: 10.1093/biomethods/bpx006

Figure Lengend Snippet: Comparison of available tools for constructing libraries for next-generation sequencing

Article Snippet: Based on the two DNA samples tested, both 50ng and 75ng starting amounts of DNA yielded very similar results, and similarly there was no obvious benefit when less diluted pre-amplified product (2.5 versus 5-fold dilution) was used for Fluidigm Access Array multiplex PCR ( ).

Techniques: Comparison, Multiplex Assay, Control

Journal: Biology Methods & Protocols

Article Title: PrimerPooler: automated primer pooling to prepare library for targeted sequencing

doi: 10.1093/biomethods/bpx006

Figure Lengend Snippet: Coverage of targeted sequence regions by Fluidigm Access Array multiplex PCR and Illumina MiSeQ sequencing. Two FFPE DNA samples were tested under various experimental conditions including 50- and 75-ng DNA for pre-amplication, and 1l of 2.5- and 5.0-fold diluted pre-amplified products for Fluidigm Access Array multiplex PCR. An average of 3269 reads was achieved for the targeted regions, with 95% of targets covered by at least 50 reads, the minimal depth of reads for confident variant calling.

Article Snippet: Based on the two DNA samples tested, both 50ng and 75ng starting amounts of DNA yielded very similar results, and similarly there was no obvious benefit when less diluted pre-amplified product (2.5 versus 5-fold dilution) was used for Fluidigm Access Array multiplex PCR ( ).

Techniques: Sequencing, Multiplex Assay, Amplification, Variant Assay

Journal: The Journal of Molecular Diagnostics : JMD

Article Title: Somatic Mutation Screening Using Archival Formalin-Fixed, Paraffin-Embedded Tissues by Fluidigm Multiplex PCR and Illumina Sequencing

doi: 10.1016/j.jmoldx.2015.04.008

Figure Lengend Snippet: Outline of experimental design for mutation screening using Fluidigm Access Array PCR and Illumina MiSeq sequencing. FFPE, formalin-fixed, paraffin-embedded; QC, quality control.

Article Snippet: Experimentally, we established a practical protocol with various quality control steps for multiplex PCR with the Fluidigm Access Array, providing a uniform amplification of target genes for Illumina MiSeq sequencing.

Techniques: Mutagenesis, Sequencing, Formalin-fixed Paraffin-Embedded

Journal: The Journal of Molecular Diagnostics : JMD

Article Title: Somatic Mutation Screening Using Archival Formalin-Fixed, Paraffin-Embedded Tissues by Fluidigm Multiplex PCR and Illumina Sequencing

doi: 10.1016/j.jmoldx.2015.04.008

Figure Lengend Snippet: Strategies for development and improvement of in-house variant calling algorithm. At first, the performance of the in-house variant calling algorithm was assessed and tuned against 114 known mutations by Sanger sequencing. The additional novel variants identified by Fluidigm PCR/MiSeq sequencing were further validated by PCR and Sanger sequencing, where necessary by cloning and sequencing of the PCR products. The resulting sequence data were used to further fine-tune the algorithm. Taken together, a total of 159 Sanger sequencing confirmed somatic mutations, including 147 substitutions and 12 indels (range, 1 to 33 bp) were used to optimize the algorithm.

Article Snippet: Experimentally, we established a practical protocol with various quality control steps for multiplex PCR with the Fluidigm Access Array, providing a uniform amplification of target genes for Illumina MiSeq sequencing.

Techniques: Variant Assay, Sequencing, Clone Assay

Journal: The Journal of Molecular Diagnostics : JMD

Article Title: Somatic Mutation Screening Using Archival Formalin-Fixed, Paraffin-Embedded Tissues by Fluidigm Multiplex PCR and Illumina Sequencing

doi: 10.1016/j.jmoldx.2015.04.008

Figure Lengend Snippet: Impact of DNA quality on baseline sequence errors by Fluidigm Access Array PCR and Illumina MiSeq sequencing. A: The level of both reproducible (seen in both replicates) and nonreproducible (seen only in one replicate) variants according to alternate allele frequency (AAF) and DNA quality. Formalin-fixed, paraffin-embedded (FFPE) DNA samples are further divided into subgroups according to the size of genomic sequence amenable for amplification by conventional multiplex quality control PCR. The data from FFPE tissue DNA are based on 142 cases investigated by two sets of independent experiments (Figure 4) and are shown from one set of the experiments. The levels of both reproducible and nonreproducible variants, particularly the latter, are remarkably high toward lower AAF. Because of small numbers of data points with an AAF >5%, these data are combined and presented in groups as indicated. B: The percentage of reproducible variants of the total (reproducible plus nonreproducible variants) according to AAF and DNA quality as above. Nonreproducible variants are baseline sequence errors. Reproducible variants at high AAFs are likely true mutations, but those at lower AAFs are probably a mixture of false-positive and subclonal genetic changes. Thus, the proportion of reproducible variants could indicate the level of background noise and a putative threshold level of AAF to be used for detection of somatic mutation. The percentage of reproducible variants critically depends on AAF and DNA quality, being minimal at lower AAF, but increasing steadily at >10% AAF in high molecular weight (HMW) and FFPE tissue DNA samples amenable for PCR of ≥300-bp genomic fragments but not in those only supporting PCR of up to 200 bp. C: Quantification of functional copy number of DNA by TaqMan real-time PCR. The level of functional copies amenable to PCR critically depends on DNA quality, being 80% in HMW DNA but only approximately 1% in DNA samples amplifiable for PCR of up to 200 bp.

Article Snippet: Experimentally, we established a practical protocol with various quality control steps for multiplex PCR with the Fluidigm Access Array, providing a uniform amplification of target genes for Illumina MiSeq sequencing.

Techniques: Sequencing, Formalin-fixed Paraffin-Embedded, Amplification, Multiplex Assay, Mutagenesis, Molecular Weight, Functional Assay, Real-time Polymerase Chain Reaction